Veterinary World, EISSN: 2231-0916 RESEARCH ARTICLE Available at www.veterinaryworld.org/Vol.14/March-2021/1.pdf Open Access

The vulnerary potential of botanical medicines in the treatment of bacterial pathologies in fish

Farida Nurzhanova1 , Gaisa Absatirov1 , Bekzhasar Sidikhov1 , Alexander Sidorchuk2 , Nurbek Ginayatov1 and Kenzhebek Murzabaev1

1. Zhangir Khan West Agrarian-Technical University, Republic of Kazakhstan; 2. Moscow State Academy of Veterinary Medicine and Biotechnology named after K.I. Skryabin, Russia. Corresponding author: Farida Nurzhanova, e-mail: [email protected], Co-authors: GA: [email protected], BS: [email protected], AS: [email protected], NG: [email protected], KM: [email protected] Received: 06-09-2020, Accepted: 15-01-2021, Published online: 02-03-2021 doi: www.doi.org/10.14202/vetworld.2021.551-557 How to cite this article: Nurzhanova F, Absatirov G, Sidikhov B, Sidorchuk A, Ginayatov N, Murzabaev K (2021) The vulnerary potential of botanical medicines in the treatment of bacterial pathologies in fish, Veterinary World, 14(3): 551-557.

Abstract Background and Aim: The use of -based medicine in treating and preventing fish disease has become increasingly popular due to the resistance of bacterial pathogens to chemicals widely used in aquaculture. This study explored the vulnerary effect of botanical medicines made from local raw materials (greater celandine [ majus L.], St. John’s wort [ L.], and bur beggar-ticks [Bidens tripartita L.]) in the treatment of sturgeon bacterial pathologies in a controlled environment. Materials and Methods: The vulnerary activity of herbal infusions was studied on spontaneously infected fish by assessing the degree of wound healing at regular intervals: The state of the wound, reduction of the wound surface area, the formation of granulation tissue, epithelization, and wound contraction. Results: A positive vulnerary effect of C. majus, H. perforatum, and B. tripartita was observed, consistent with the use of these in folk and traditional medicine. The plant materials eliminated infection, had anti-inflammatory and vulnerary effects, stimulated granulation tissue development, and enhanced regeneration. Compared with widely accepted methods (antibiotics and other chemotherapeutic agents), botanical medicine facilitated more effective treatment over the same period without side effects. Conclusion: Practical use and the results of this study show the potential of using herbal infusions for therapeutic purposes in aquaculture. Keywords: fish, plant raw material, vulnerary effect, wound. Introduction botanical therapies in fish [8-21]. Medicinal plants are The aquaculture industry has experienced an used as immunostimulant, stress-relieving [10-12], emergence of diseases of various etiologies due to high antiparasitic [13,14], antioxidant, and antimicrobial stocking density and stressful conditions. Among the agents [15-21]. pathologies that occur when keeping fish in recirculat- Plant extracts of Boesenbergia pandurata, ing aquaculture systems, the most common are bacte- Salmo ferox, and Zingiber zerumbet for the preven- rial diseases caused by Aeromonas and Pseudomonas tion of illness caused by Aeromonas hydrophila species [1-7]. and Pseudomonas spp. have a positive effect on the Fish disease treatment strategies are based pri- non-specific immune response and inhibit bacterial marily on the use of antibiotics and chemotherapy. infection [18]. Bioactive compounds of Origanum vul- However, in recent years, plant-based medicines have gare have immunostimulant, cytotoxic, bactericidal, become increasingly popular in aquaculture due to the and antioxidant properties, which make them promis- resistance of bacterial pathogens to standard antibiot- ing for fish aquaculture [19]. A study of B. pandurata, ics, the accumulation of antibiotics in food fish, and S. ferox, and Z. zerumbet showed their antibacterial environmental pollution. properties in preventing and treating infections caused Studies of the past decades showed encour- by A. hydrophila and Pseudomonas fluorescens [20]. aging results and opened up new opportunities for Studies of the immunostimulant effects of brier (Rosa canina) on the hematological and immune parameters Copyright: Nurzhanova, et al. Open Access. This article is of sturgeons infected with Mycobacterium salmoniph- distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/ ilum showed a non-specific immune response and the by/4.0/), which permits unrestricted use, distribution, and antibacterial effect of this plant [21]. reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the In addition to a wide range of anti-inflamma- Creative Commons license, and indicate if changes were made. tory, antimicrobial, and immunostimulant properties, The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data biologically active plant components also exhibit vul- made available in this article, unless otherwise stated. nerary activity [22-29]. Plants and their extracts have

Veterinary World, EISSN: 2231-0916 551 Available at www.veterinaryworld.org/Vol.14/March-2021/1.pdf great potential for the treatment of pathogenic and feed contained 20.03 MJ of assimilated energy and non-pathogenic wounds. 47.0% crude protein. Feeding was carried out by hand Data on the potential for wound healing of plant twice a day according to the calculated rates. extracts used in aquaculture are scarce. There are stud- Experimental models ies on the wound healing effect of walnut leaves and To study wound healing, we used fish sponta- onion leaves on wounds in Clarias gariepinus [30], neously (naturally) affected by pseudomonosis and accelerated wound healing in African catfish when aeromonosis, with moderate disease severity. Sturgeon Ocimum gratissimum was added to the feed [31], individuals with pronounced clinical signs of a bacte- and wound healing in Labeo rohita fish with cur- rial disease were identified during examination. Based cumin [32]. Healing in fish has mainly been studied on the clinical examination results, bacteriological in artificially modeled wounds or mechanical inju- studies were performed to confirm the preliminary ries. Little is known about the effect of plant extracts diagnoses of pseudomonosis and aeromonosis. on the healing of wounds caused by pathogenic Typical bacterial septicemia with ulceration and fin microorganisms. erosion, hemorrhagic ulcerative lesions on the skin (both This study explored the vulnerary effect of small on the skin surface and penetrating deep into the botanical medicines made from local raw materials muscles) were observed in sturgeons infected with pseu- (greater celandine [Chelidonium majus L.], St. John’s domonosis. Hemorrhagic septicemia was noted in fish wort [Hypericum perforatum L.], and bur beggar-ticks infected with aeromonosis, characterized by abdominal [Bidens tripartita L.]) in the treatment of sturgeon distention and ascitic fluid accumulation, small superfi- bacterial pathologies in a controlled environment. cial lesions, and local hemorrhage in the gills. Materials and Methods Solution preparation Ethical approval A stock solution was prepared from each plant The experimental plan and procedures with material at a dilution of 1:20. Crushed plant mate- fish within the framework of the current research rial (50 g) was added to 1000 mL distilled water and were approved by the local commission on biologi- infused in a boiling water bath for 30 min. The infu- cal ethics of TOO Batys Zoo Vet Servis (protocol No. sion was cooled, filtered, and used in therapeutic baths 12 dated 01/10/2018). The experiments were con- with an exposure of 30 min with a dilution of 500 mL ducted according to the recommendations set out in of the initial solution per 50 L water. the “Rules of Good Clinical Practice of the Eurasian Experimental design Economic Union” N 79, 03.11.2016. Three treatment groups were prepared: Group Study location and period 1 (n=15, 53% with pseudomonosis, 47% with aero- The study was conducted from September 2017 monosis; C. majus infusion); Group 2 (n=15, 67% with to January 2020 in the RAS at the “Educational pseudomonosis, 33% with aeromonosis; B. tripartita and Scientific Complex of the Pilot Production of infusion); Group 3 (n=15, 60% with pseudomonosis, Aquaculture” LLP, where various species of sturgeon 40% with aeromonosis; H. perforatum infusion), and are grown in Uralsk, Republic of Kazakhstan. a control group (n=15, 53% with pseudomonosis, Plant material 47% with aeromonosis) were formed using the analog The following medicinal plants were selected method. Fish in each experimental group were treated as raw materials for the botanical medicines used with herbal infusions every other day. The control in the study: Greater celandine (Chelidonium group was not treated. majus L.) [24,28], St. John’s wort (Hypericum perfor- Wound healing intensity atum L.) [28,29], and bur beggar-ticks (Bidens tripar- The healing process was observed by taking pho- tita L.) [27,28]. tographs, starting from the day the experiments began. Experimental conditions The wound healing properties of the test plants were The fish were kept in quarantine pools. During the evaluated clinically (the state of the wound, reduc- 2 experiment, the temperature in the closed water supply tion of the wound surface area (on average, mm ), unit was maintained at an optimal level of 20-23°C. the formation of granulation tissue, epithelization, The main parameters of the aquatic environment and wound contraction) at regular intervals of 3, 5, 7, corresponded to fishery standards: pH 7.0-8.0; dis- 10, and 15-20 days from the beginning of the experi- solved oxygen 4.0 mg/L; permanganate oxidizability ment. Wounds were considered healed if the colliqua- tive granulation tissue was no longer visible and the 10.0 mgO2/L; ammonia (NH4+) 0.5 mg/L; total hard- ness 6.0-8.0 mg/L; and biochemical oxygen demand wound was covered with a new epithelium. Survival during the experiment was taken into account. 5 2.0 mgO2/L. When feeding the fish, an extruded compound feed was used, produced by the extrusion Statistical analysis method, and consisting of fish meal (57.5%), soybean Digital data processing was carried out using meal (20.0%), wheat grain (1.5%), fish oil (20.0%), the method of variation statistics using Microsoft and premix (1.0%). One kg sample of compound Excel 2007.

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Results and Discussion During treatment on the 3rd and 5th days of the Clinical performance at the beginning of the exper- experiment, a decrease in inflammation and puffi- iment (day 0) in all experimental groups was as follows: ness at the edge of the wound was observed, and the Experimenters marked single or multiple ulcers of vari- first signs of healthy granulation tissue appeared in ous surface areas and depths deep in the muscles with the the form of separate focal areas. Eventually, it filled formation of rims of bright red color along the edges on up the entire wound in all three experimental groups the dorsal, abdominal, and tail parts of the fish. Redness (Figures-1b and c-3b and c). of the abdomen and the area around the anus was noted. Subsequently, the transition of granulation tissue Severe hyperemia and puffiness of the skin tis- from bright red to pink characterized the proliferation sue were observed at the verge of the wound surface and formation of the epithelium, which grew from (Figures-1a-3a). the periphery to the center of the wound, covering the

a b c

d e f Figure-1: Macroscopic changes in the wound in the first experimental group (treatment with Chelidonium majus): (a) At the beginning of the experiments; (b) the 3rd day of the experiments; (c) the 5th day of the experiments; (d) the 7th day of the experiments; (e) the 10th day of the experiments; (f) the 15th day of the experiments.

a b c

d e f Figure-2: Macroscopic changes in the wound in the second experimental group (treatment with Bidens tripartite): (a) At the beginning of the experiments; (b) the 3rd day of the experiments; (c) the 5th day of the experiments; (d) the 7th day of the experiments; (e) the 10th day of the experiments; (f) the 15th day of the experiments.

Veterinary World, EISSN: 2231-0916 553 Available at www.veterinaryworld.org/Vol.14/March-2021/1.pdf wound bed, which indicated the beginning of wound worsening illness, feed refusal, and an increase in the shrinkage. The growth of granulation tissue led to external symptoms of the disease were observed. decreased wound depth, and epithelial cell growth led The wound process transition from the formation to a decreased wound area. of granulation tissue to epithelization and regeneration In the control group, inflammation and puffi- was observed in all three experimental groups, indicat- ness at the edge of the wound surface did not resolve, ing the effectiveness of the herbal infusions. However, and excessive mucoid discharge was noted. In most the treatment effect on the change in wound surface cases, signs of necrosis and sloughing developed, and area was significantly more pronounced in C. majus ulcers became larger in several individuals. There and H. perforatum groups, respectively (Table-2). was no reduction in size or contraction of the wound In the study of wound healing dynamics on the (Figures-4a and b). Fish died in the case of transition surface of the body of sick fish, a change in wound to a generalized form (Table-1). surface area was observed in the first and third exper- Table-1 shows that eight fish died in the control imental groups compared to the second experimental group, and the survival rate was 46%. All fish sur- and control groups. Starting from the 7th day after the vived (100% survival) in the first and third experi- beginning of the experiments, the groups treated with mental groups. One fish died (93% survival) in the celandine and St. John’s wort showed a substantial second group. reduction in wound size. The decrease in the wound During the experiment, an improvement in the surface area was 41.8% in Group 1 and 55.2% in general condition of the fish and the clinical signs Group 3. Subsequently and throughout the observation of disease disappeared in all experimental groups. period (up to 20 days), the wound surface area in these The fish willingly ate the feed. In the control group, groups continued to decrease (Figures-1d-f and 2d-f).

a b c

d e f Figure-3: Macroscopic changes in the wound in the second experimental group (treatment with Hypericum perforatum): (a) At the beginning of the experiments; (b) the 3rd day of the experiments; (c) the 5th day of the experiments; (d) the 7th day of the experiments; (e) the 10th day of the experiments; (f) the 15th day of the experiments.

Table-1: Fish survival during the experiment.

Groups The period of the experiment (days) Survival rate, % 3rd day 5th day 7th day 10th day 15th day 20th day Fish mortality First group ‑ ‑ ‑ ‑ ‑ ‑ 100 (n=15) Second group 1 ‑ ‑ ‑ ‑ ‑ 93 (n=15) Third group ‑ ‑ ‑ ‑ ‑ ‑ 100 (n=15) Control group 4 2 1 1 ‑ ‑ 46 (n=15)

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Table-2: The dynamics of wound healing in fish.

Groups The average area of the wound (mm) Wound healing, % Start 3rd day 5th day 7th day 10th day 15th day 20th day First group 28.72±6.22 27.20±5.84** 23.91±5.09** 16.71±4.17 9.55±3.09 4.79±1.71 1.89±0.73 93.4 (n=15) Second group 20.39±4.37 17.23±4.24 16.24±4.03 15.77±4.00 15.33±3.93 14.86±3.93 14.39±3.91 29.5 (n=15) Third group 25.00±5.80 20.15±5.26 15.48±4.45 11.12±3.38 7.43±2.50 3.80±1.47 1.67±0.65 93.3 (n=15) Control group 19.03±4.31 11.47±3.15 8.60±2.93 8.24±3.20 7.57±3.24 7.84±3.33 8.19±3.58 0 (n=15) *p<0.05; **p<0.01; ***p<0.001 in comparison with the control

The maximum values by the 15th and 20th days of the experiments were found in Groups 1 and 3 (4.79±1.71 mm2 [83.3%] and 1.89±0.73 mm2 [93.4%]; and 3.80±1.47 mm2 [84.8%] and 1.67±0.65 mm2 [93.3%], respectively). It should be noted that at all times during the experiments, although there was no pronounced effect on wound size in Group 2, the effect of the medicinal a b infusion on the healing process was similar to wound Figure-4: A wound in the control group (without healing in the groups treated with celandine and St. treatment): (a) At the beginning of the experiment; (b) at John’s wort: A marked decrease in inflammation, the the end of the experiment. recession of puffiness, intensive formation of granu- lation tissue, and epithelization of wounds with slight scar formation toward the center of the wound, which was almost completed by the end of the experiments (Figures-3d-f). The wounds were closed with an epi- thelium with a relatively small reduction in area. Therefore, bur beggar-ticks have a vulnerary effect, as do celandine and St. John’s wort, which con- tribute to the healing process. Figure-5 shows that the reduction in wound size in the first and third groups was significant compared with the second and control groups, which indicated intensive healing. Figure-5: The dynamics of wound size reduction, mm2. A visual assessment of wounds showed a decrease in inflammatory processes, granulation tissue growth, better than bur beggar-tick and control. Bidens tripar- epithelial growth, tissue regeneration, and remolding of the tissues in the first, second, and third groups. tite infusion and C. majus and H. perforatum infusions The site of the wound remained slightly darker than had a healing effect on the wound but did not signifi- the skin. Wound size decreased with the formation of cantly affect wound size. granulation tissue. No cases of recurrent disease were registered The reduction of the wound in the first and third during the observation period for fish from the exper- groups began by the 3rd and 5th day with granulation imental groups. tissue formation and the formation and migration of In the available literature, there is a significant epithelial cells from the edges to the center of the amount of scientific research on the use of medicinal wound. A slight reduction in the wound size was plants as immunostimulants that increase disease resis- observed in the second group with a pronounced vul- tance in fish by enhancing their non-specific and spe- nerary process. cific immunity [15-21]. However, there are very few Compared to the control group, the vulnerary studies on the wound healing effect of herbal drugs on process was more pronounced in the groups treated spontaneously emerging wound pathologies of bac- with C. majus, H. perforatum, and B. tripartita L. The terial etiology in fish under real-world conditions in wound size reduction was more pronounced in the an aquarium complex. There are separate studies on first and third groups than in the second and control herbal drugs for artificially simulated wound patholo- groups. gies in fish [31-33]. The results demonstrate that C. majus and H. per- Our research indicates that the proposed types foratum infusions reduced the surface area of wounds and forms of herbal preparations, characterized by

Veterinary World, EISSN: 2231-0916 555 Available at www.veterinaryworld.org/Vol.14/March-2021/1.pdf bacteriostatic and bactericidal action, promote the Industrial Production of Aquaculture” LLP for assis- activation of non-specific immune mechanisms and tance in conducting experimental work. The authors have a pronounced wound healing effect. Herbal did not receive financial support for the study. preparations, unlike chemotherapeutic ones, are eco- Competing Interests logically safe and do not adversely affect the quality and nutritional value of fish products. These findings The authors declare that they have no competing will be useful for the development of environmen- interests. tally friendly strategies for controlling fish disease in Publisher’s Note aquaculture. Veterinary World remains neutral with regard Conclusion to jurisdictional claims in published institutional This study showed a positive vulnerary effect affiliation. of C. majus, H. perforatum, and B. tripartita, consis- References tent with the use of these plants in folk medicine and 1. Kazarnikova, A.V. and Shestakovskaya, E.V. (2005) Major traditional medicine. The use of raw plant material as sturgeon diseases in aquaculture. Publishing House of a therapeutic agent eliminated infection, had anti-in- VNIRO, Russian Federation Research Institute of Fishery flammatory and vulnerary effects, stimulated the and Oceanography, Moscow. p104. development of granulation tissue, and enhanced the 2. Kazarnikova, A.V. and Shestakovskaya, E.V. (2005) Diseases of Sturgeon Fish During Artificial Reproduction regeneration process due to the content of biologically and Commercial Cultivation. Publishing House Kola active components in medicinal plants. Each biolog- Science Centre, Apatity. p58. ically active agent may have a specific function and 3. Ginayatov, N.S., Zalyalov, I.N. and Absatirov, G.G. (2016) affect one or more phases of the vulnerary process. Identification of the pathogen of infectious pathology of A comparative study of the vulnerary properties sturgeon fish in a recirculating aquaculture system. Kazan State Acad. Vet. Med., 2(226): 42-45. of C. majus, H. perforatum, and Bidens tripartite with 4. Nurzhanova, F.K., Absatirov, G.G., and Ezhkova, M.S. bacterial pathologies of sturgeon showed that infu- (2018) Epizootic State of Sturgeon when Bred in a sions of C. majus and H. perforatum were the most Recirculating Aquaculture System. III National Scientific promising. Bidens tripartite infusion also had a heal- and Practical Conference Status and Paths of Aquaculture Development in the Russian Federation in the Light ing effect on the wound, but no noticeable effect on of Import Substitution and Ensuring Food Security of reducing the wound size was observed; however, we the Country. Amirit Technologies, Inc., Kazan, Russia. recommend using this herbal decoction as an auxil- p203-207. iary substance in complex therapeutic and prophylac- 5. Ture, M., Ozcelep, T., Akbulut, B. and Kutlu, I. (2018) tic measures. Disease of Russian sturgeon (Acipenser gueldenstaed- tii) caused by Aeromonas sp. Genet. Aquat. Organ., 2(2): Compared with traditional methods (antibiotics 43-47. and other chemotherapy methods), botanical therapy 6. Kayiş, Ş., Er, A., Kangel, P. and Kurtoğlu, İ.Z. (2017) allows treatment to be carried out more effectively Bacterial pathogens and health problems of Acipenser and as quickly as standard therapeutics but with- gueldenstaedtii and Acipenser baerii sturgeons reared in the eastern Black Sea region of . . J. Vet. Res., 18(1): out unwanted side effects. The practical use and the 18-24. results of this study show the potential of using herbal 7. Santi, M., Pastorino, P., Foglini, C., Righetti, M., Pedron, C. infusions as effective, safe, and affordable means for and Prearo, M. (2018) A survey of bacterial infections in therapeutic purposes in treating fish in aquaculture. sturgeon farming in Italy. J. Appl. Ichthyol., 35(1): 275-282. 8. Reverter, M., Bontemps, N., Lecchini, D., Banaigs, B. and Authors’ Contributions Sasal, P. (2014) Use of plant extracts in fish aquaculture as an alternative to chemotherapy: Current status and future FN: Acquisition of data, conducted experiments, perspectives. Aquaculture, 433 : 50-61. analysis and interpretation of data, and drafted the 9. Syahidah, A., Saad, C.R., Daud, H.M. and Abdelhadi, Y.M. manuscript. GA: Conception and design, supervised (2015) Status and potential of herbal applications in aqua- the research, and final approval. BS: Procedure for culture: A review. Iran. J. Fish. Sci., 14(1): 27-44. 10. Kurapova, T.M. and Kazimirchenko, OV. (2015) The effect conducting experiments, revision, and editing the arti- of biologically active substances of the series (Bidens tri- cle. AS: Comparison of research results and revised partita L.) on some immunophysiological and microbi- the article critically. NG: Statistical analysis and inter- ological indicators of the state of yearlings of carp. Bull. pretation of data, and drafted the manuscript. KM: Astrakhan State Tech. Univ. Fish., 2015(4): 69-75. Selection of literature, creating a reference list, anal- 11. Awad, E. and Awaad, A. (2017) Role of medicinal plants on growth performance and immune status in fish. Fish ysis, and interpretation of data. All authors have read Shellfish Immun., 67: 40-54. and approved the final manuscript. 12. Chakraborty, S.B. and Hancz, C. (2011) Application of phytochemicals as immunostimulant, antipathogenic and Acknowledgments antistress agents in finfish culture. Rev. Aquaculture, 3(3): The authors express their gratitude to the Zhangir 103-119. 13. Yao, J.Y., Zhou, Z.M., Pan, X.Y., Hao, G.J., Li, X.L., Xu, Y., Khan West Kazakhstan Agrarian-Technical University Shen, J.Y., Ru, H.S. and Yin, W.L. (2011) In vivo anthel- for providing the material and technical base and the mintic activity of from Chelidonium majus L. “Educational and Scientific Complex of Experimental against Dactylogyrus intermedius in Carassius auratus.

Veterinary World, EISSN: 2231-0916 556 Available at www.veterinaryworld.org/Vol.14/March-2021/1.pdf

Parasitol. Res., 109(5): 1465-1469. on the basis of thyme, chestnut, licorice, nettle, St. John’s 14. Degtyarik, S.M., Grebneva, E.I., Slobodnitskaya, G.V., wort. Sci. Proc. Belgorod State Univ. Ser. Med. Pharm., Benetskaya, N.A., Bespalyi, A.V., and Maksimyuk, A.V. 10-2(129): 114-117. (2017) Phytopreparation for the Treatment and Prevention 23. Zhelonkin, N.N., Pervushkin, S.V., Klimova, L.D., Ber, O.V., of Trichodiniosis in Sturgeon Fish. Aquaculture of Sturgeon. Sokhina, A.A. and Budanova, A.S. (2015) Development of Challenges and Prospects. Сollected Papers of Articles of the a combined medicinal form of spray for the treatment of International Scientific-Practical Conference. Publishing ecologically caused dermatitis. Izvestia Samara Sci. Center House “Astrakhan University”, Astrakhan, Russia. p77-79. Russ. Acad. Sci., 5-2(17): 579-582. 15. Degtyarik, S.M., Grebneva, E.I., Slobodnitskaya, G.V., 24. Ruzieva, I.G. and Karomatov, I.D. (2018) A promising Benetskaya, N.A., Bespalyi, A.V. and Maksimyuk, E.V. means of celandine phytotherapy. Biol. Integr. Med., 2(19): (2016) The Effect of Plant Extracts on the Causative Agents 77-86. of Aeromonosis and Pseudomonosis of Fish. Collection 25. Dutova, O.G. and Petrenko, V.A. (2019) Bacteriological of Research Papers Issues of Fisheries in Belarus, Minsk. control of the state of the wound microflora in the treat- p249-261. ment of skin wounds with extracts from medicinal plant raw 16. Degtyarik, S.M., Grebneva, E.I., Slobodnitskaya, G.V., materials. Bull. Altai State Agrarian Univ., 1(171): 73-77. Benetskaya, N.A., Bespalyi, A.V. and Maksimyuk, E.V. 26. Frolova, A.V., Kosinets, A.N., Zholnerovich, M.L. and (2017) The effect of plant phytoncides on the viability and Grushin, V.N. (2009) Experimental selection of medicinal virulence of etiological agents of bacterial infections in fish. plant origin for local treatment of purulent wounds. Proc. Bull. Natl. Acad. Sci. Belarus Agrarian Sci. Ser., 2017(1): Vitebsk State Acad. Vet. Med., 45(1): 99-103. 79-89. 27. Karomatov, I.D. and Abduvokhidov, A.T. (2017) The 17. Tkachenko, G.M., Buyun, L.I., Terekh-Maevskaya, E. and famous medicinal plant bur beggar-ticks. Biol. Integr. Med., Osadowski, Z. (2017) Screening for antimicrobial activity 9 : 12-22. of ethanol extract isolated from the leaves of Ficus hispida 28. Vasfilova, E.S., Tretyakova, A.S., Podgaevskaya, E.N., l.f. (Moraceae) relative to fish pathogens. Trans. Far East. Zolotareva, N.V., Khokhlova, M.G., Igosheva, N.I., State Tech. Fish. Univ., 41(3): 56-62. Ektova, S.N., Morozova, L.M. and Mukhin, V.A. (2014) 18. Hardi, E.H., Kusuma, I.W., Suwinarti, W., Wild Medicinal Plants of the Urals. Tutorial. Publishing Saptiani, G., Sumoharjo, and Lusiastuti, A.M. (2017) house Ural University, Ekaterinburg. p204. Utilization of several herbal plant extracts on Nile tilapia 29. Avdachenok, V.D. and Tuminets, O.A. (2017) The Use of in preventing Aeromonas hydrophila and Pseudomonas sp. Hypericum perforatum in Mixed Infections in Ruminants. bacterial infection. Nusantara Biosci., 9(2): 220-228. Guidelines. Vitebsk State Academy of Veterinary Medicine, 19. Beltrán, J.M.G., Espinosa, C., Guardiola, F.A. and Vitebsk. p12. Estebana, M.A. (2018) In vitro effects of Origanum vulgare 30. Bello, O.S., Emikpe, B.O., Olaifa, A.K. and Olaifa, F.E. leaf extracts on gilthead seabream (Sparus aurata L.) leuco- (2013) Investigation into the healing properties of walnut cytes, cytotoxic, bactericidal and antioxidant activities. Fish (Tetracarpidium conophorum) leaf and onion (Allium cepa) Shellfish Immun., 79: 1-10. bulb residues in Clarias gariepinus. Arch. Med. Vet., 45(3): 20. Hardi, E.H., Nugroho, R.A., Kusuma, I.W., Suwinarti, W., 291-297. Sudaryono, A. and Rostika, R. (2019) Borneo herbal 31. Abdel-Tawwab, M., Emikpе, B., Adeshina, I., Ajani, E.K., plant extracts as a natural medication for prophylaxis and Jenyo-Oni, A. and Tiamiyu, L.O. (2019) Effect of dietary treatment of Aeromonas hydrophila and Pseudomonas clove basil, Ocimum gratissimum, leaves extract on healing fluorescens infection in tilapia (Oreochromis niloticus). of artificially wounded African catfish, Clarias gariepinus F1000Research, 7(1847): 1-18. (B.), juveniles. J. Appl. Aquaculture, 31(4): 289-300. 21. Duman, S. and Şahan, A. (2018) Some hematological and 32. Kumari, U., Verma, N., Nigam, A.K., Mittal, S. and non-specific immune responses of rosehip (Rosa canina) - Mittal, A.K. (2016) Wound-healing potential of curcumin in Fed Russian sturgeon (Acipenser gueldenstaedtii Brandt & the carp, Labeo rohita. Aquaculture Res., 48(5): 2411-2427. Ratzeburg, 1833) to Mycobacterium salmoniphilum. Braz. 33. Verma, N., Kumari, U., Mittal S. and Mittal, A.K. (2017) Arch. Biol. Technol., 61(14): e18180283. Effect of asiaticoside on the healing of skin wounds in the 22. Volodina, T.A., Penevskaya, N.A. and Stepanova, E.F. carp Cirrhinus mrigala: An immunohistochemical investi- (2012) Development and research of phytopreparation gation. Tissue Cell, 49(6): 734-745. ********

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